Adenine phosphoribosyltransferase (APRT), an enzyme of purine salvage, utilizes adenine and 5-phosphoribosyl-1-pyrophosphate to produce adenosine monophosphate and pyrophosphate. In man, complete APRT deficiency is a supposedly rare inborn error of purine metabolism that is inherited in an autosomal recessive manner. Adenine, which is primarily a byproduct of polyamine biosynthesis, accumulates to high levels in APRT deficient individuals and is ultimately oxidized to 2,8-dihydrixyadenine (DHA). Although the defect can be relatively benign, DHA is nephrotoxic and can lead to life-threatening DHA urolithiasis. A relatively high frequency of apparent heterozygosity for this disorder suggests that homozygosity could be more frequent than is currently recognized, presumably due to highly variable clinical expression coupled with problems of diagnosis. We have cloned a functional human APRT gene and have determined the nucleotide sequence of its coding regions and introns. We also have cell cultures from APRT deficient patients and APRT deficient human cell clones obtained by selection from normal somatic cells in vitro. Using our cloned gene to probe these cells, we will determine the molecular bases for expression of human APRT deficiency. By comparing mutant APRT genes in the human population to those obtained from cultured, human somatic cells we will ascertain whether or not somatic cell mutation in vitro, which forms the basis of most mutagenesis assays, is qualitatively different from mutation in germ cells in situ. Mutant cell nucleic acids will be analyzed by Southern and northern blots, RNase A digestion of DNA/RNA molecular hybrids and DNA sequence analysis. Mutant APRT proteins will also be analyzed using antisera directed against the N or C-termini of the normal enzyme.